GSM6620594: s030714; Mus musculus; RNA-Seq
Single cells were randomly picked into precooled lysate by mouth. After shaking for 1min, the lysates were incubated at 72℃ for 3min to release RNA and initial denaturation. Herein, STRT-seq (modified single-cell tagged reverse transcription protocol) was applied. Firstly, single cells are picked into a PCR tube containing lysates, oligo(dT) primers, and barcode primers. After reverse transcription and cDNA amplification, the single cells labelled with different barcodes were pooled together for subsequent library construction. Additional amplifying for pooled cDNAs were performed by biotin-indexed primers, following shearing to about 300bp by sonication Then, Dynabeads® MyOne™ Streptavidin C1 beads were used to enrich for the 3' terminal of cDNAs. Finally, the library was constructed by Kapa Hyper Prep Kit and sequenced on illumina Hiseq X Ten for 150bp pair-end reads.